Figure 5 | Scientific Reports

Figure 5

From: SgRNA Expression of CRIPSR-Cas9 System Based on MiRNA Polycistrons as a Versatile Tool to Manipulate Multiple and Tissue-Specific Genome Editing

Figure 5

MiRNA based sgRNA cassettes can be modified to alter the proportion of HR in genome editing. (A) Schematic overview of pcDNA3.1(−) shNHEJ-TLR1.1 sgRNA construction. (B) Schematic overview of the principle of detection of the ratio of HR and NHEJ. qRT-PCR (C) and luciferase reporter assay (D) were used to demonstrate that Lig4 was effectively suppressed by pcDNA3.1(−) shNHEJ TLR1.1 sgRNA. (E) Gene editing with pcDNA3.1 (−) shNHEJ TLR1.1 sgRNA had a higher efficiency of homologous recombination compared to pLVX-u6-TLR1.1 sgRNA. pcDNA3.1(−) shNHEJ TLR1.1 sgRNA or corresponding control was co-transfected into 293T with Cas9 expression plasmid, DSB repair reporter construction, and corresponding donor. Afterwards, luciferase assay was performed 3 days after transfection.

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